Abstract Many adult tissues replace cells lost due to normal wear and tear via the activity of stem cells, but can use entirely different strategies when injury occurs. The ability to understand and control regeneration, or the regrowth of lost tissues or organs in response to injury, is a long-standing goal in biology. Cells with the capacity to adopt the biological properties of other cell types under specific conditions, or cellular plasticity, are key contributors to regeneration. Stem cells and even differentiated cells can have surprising degrees of plasticity, allowing them to adopt new fates and rebuild damaged tissues. This happens in response to altered microenvironments that arise upon injury, but the mechanisms that regulate plasticity are poorly understood. In the prior funding period, we showed that a damaged organ converts non-stem cells to stem cells to repair the tissue. Since cellular plasticity is emerging as a general feature of tissue regeneration, we identify mechanisms regulating the regeneration of adult somatic cells in vivo using two functionally similar systems: mammalian and Drosophila testes. In Aim 1 we build on our finding that quiescent somatic cells in the Drosophila testis transdifferentiate into stem cells upon damage, using live imaging, lineage tracing and genetic manipulation to identify the regulatory genes. We also study the aberrant consequence of regeneration, ectopic stem cell niche formation in Drosophila testes, and obtain gene expression data from rare cells in this tissue to piece together the mechanisms that control this process. In Aim 2 we extend our new preliminary data showing that damage to the adult mouse testis, in the form of ablation of Leydig cells, triggers regeneration of these important quiescent steroidogenic cells. We characterize cells giving rise to new Leydig cells upon damage and aging at the cellular and molecular level. Together these Aims will reveal the molecular cues regulating loss of quiescence and the induction of regeneration within two different systems in vivo, providing a potent means to determine the mechanisms sustaining damage-induced tissue repair.